Reduced intracellular pH in lymphocytes from the spontaneously hypertensive rat

This study was designed to determine the cytoplasmic pH (pH_i) profile of lymphocytes from a rat model of genetic hypertension that is well suited for study before and after the development of spontaneous hypertension. For this purpose, pH, was measured in thymic lymphocytes obtained from spontaneously hypertensive rats (SHR) and from age-matched Wistar-Kyoto (WKY) control rats using 2′,7′-bis carboxyethyl-5,6-carboxyfluorescein (BCECF), a pH-sensitive fluorescence probe. At the age of 16-20 weeks, pH_i of lymphocytes suspended in a HCO₃-free HEPES-buffered solution, was markedly lower in the SHR than in the WKY rats (7.07±0.02, n=16 and 7.22±0.01, n=15, respectively, p<0.001), whereas systolic blood pressure was higher in SHR than in WKY rats (175±5.0 and 105±3.0 mm Hg, respectively, p<0.001). In rats less than 5 weeks of age, pH, was also lower in SHR than in WKY rat lymphocytes (7.12±0.04, n=11 and 7.23±0.04, n=11, respectively, p<0.05), although at this age systolic blood pressure was not different between the two groups (87±4.0 and 85±3.0 mm Hg, respectively). In lymphocytes suspended in a more physiological HCO₃/CO₂-buffered solution, pH_i was again lower in the adult SHR than in the WKY rat (7.18±0.02, n=16 and 7.31±0.02, n=16, respectively, p<0.001). Under this condition, blockade of Na⁺-H⁺ exchange by a specific inhibitor, ethyl-isopropyl-amiloride (EIPA), or by exposure to Na⁺-free media resulted in intracellular acidification in both SHR and WKY rat lymphocytes. The difference in pH_i between SHR and WKY rat lymphocytes persisted after blockade of the Na⁺-H⁺ exchanger by either EIPA (7.09±0.04 vs. 7.20±0.03, p<0.05) or by removal of external Na⁺ (7.04±0.05 vs. 7.19±0.02, p<0.005). Our data show that reduced pH, is a feature of lymphocytes from the SHR that is apparent before and after the development of hypertension. Persistence of reduced pH, in SHR cells, as compared with WKY rat cells, after Na⁺-H⁺ blockade suggests that a primary abnormality in the activity of the Na⁺-H⁺ exchanger is unlikely to account for the observed reduction in steady-state pH: of SHR cells. This alteration in pH_i regulation persisted after removal of external Na⁺ and was seen in the presence and in the absence of external HCO₃, indicating that a sodium-independent, bicarbonate-independent mechanism of cytoplasmic acid accumulation accounts for the observed difference in pH_i.